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Background Macrophage migration inhibitory factor (MIF) is a key pro-inflammatory cytokine,exhibiting antioxidant properties.However,the role of MIF in cardiac fibrosis is not well known.In the present study,the effects of MIF on Smad3 and Nrf2 signalings in cardiac fibroblasts were investigated.Methods Cardiac fibroblasts were isolated from 1-3 days old C57BL/6 mice,and the cardiac fibroblasts from passage 2 to 4 were used in this study.Expression of fibrosis-associated Collal,Co13al and α-SMA in mouse cardiac fibroblasts was detected by immunofluorescence staining and Weste-blot assay,respectively.Intracellular oxidants in mouse cardiac fibroblasts were measured by using the probe dichlorofluoroscindiacetate (DCFH-DA) under confocal microscopy.Weste-blot assay was also used to detect Smad3 and Nrf2,antioxidant proteins,MLL and HCF-1 in mouse cardiac fibroblasts.Results Immunofluorescence staining and Weste-blot assay showed that MIF could markedly inhibit fibrosis-associated Collal,Co13al and α-SMA expression in cardiac fibroblasts.DCFHDA staining revealed that MIF can efficiently decrease reactive oxygen species (ROS) level in Ang-Ⅱ-induced cardiac fibroblasts.Additionally,Smad3 activation was inhibited,but transcription factor Nrf2 and the downstream antioxidant genes,including HO-1,SOD-1,SOD2,Trx-2 and e-NOS,were increased in MIF-treated cardiac fibroblasts.MLL and HCF-lwere up-regulated by MIF,and either MLL knockdown or HCF-1 knockdown could consistently suppress Nrf2 expression in cardiac fibroblasts.Conclusions MIF possesses anti-fibrosis effect by inactivating Smad3 and activating Nrf2 in cardiac fibroblasts.